The purpose of this application is to delineate mechanisms which control fetal and adult hemoglobin (HbF and HbA) produced in individual erythrocytes in adults. 3 principles underlie the proposal: 1) increased HbF production can ameliorate the clinical severity of sickle cell disease, thalassemia and falciparum malaria infections; 2) in normal individuals, HbF is confined to a small minority of cells called F-cells; 3) increased HbF production after birth is primarily due to increased production of F cells. We therefore believe any future therapeutic manipulation of HbF will rquire knowledge of cell-centered mechanisms. Our specific aims are directed toward two areas of cellular regulation. First, what control the amount of HbF produced within the cell. We wish to study the interrelation between HbF and HbA within the same cell, e.g., is HbF and HbA production reciprocal in an individual red cell or is HbF added to a normal amount of HbA. To answer these questions we will develop single cell assays which can quantitate fetal and adult Hb(A or S) in the same cell. Antibodies with high affinity and high specificity are needed for this purpose and will be developed through hybridoma technology. We will also study how difference mutations affect the absolute level of HbF produced per cell. We will determine if a minimal level of HbF is necessary to insure preferential survival of F cells in sickle cell disease: Second, we have recently defined a genetic site which control F cell production and is linked to the Hb-Beta locus. The site regulates F cell levels in both non-anemic and anemic individuals. Using a variety of methods, we wish to establish whether this site is part of the GammaDeltaBeta gene complex or separate from it. We also wish to determine whether the genetic locus can be reproducibly activated in erythroid culture systems and whether this locus is in linkage disequilibrium with the betaSglobin mutation. Our studies should not only aid in an understanding of how HbF is regulated but also, through the study of F cells, we expect to learn more about the mechanisms which control cell differentiation.